Two-way communication between SecY and SecA suggests a Brownian ratchet mechanism for protein translocation

Abstract

The essential process of protein secretion is achieved by the ubiquitous Sec machinery. In prokaryotes, the drive for translocation comes from ATP hydrolysis by the cytosolic motor-protein SecA, in concert with the proton motive force (PMF). However, the mechanism through which ATP hydrolysis by SecA is coupled to directional movement through SecYEG is unclear. Here, we combine all-atom molecular dynamics (MD) simulations with single molecule FRET and biochemical assays. We show that ATP binding by SecA causes opening of the SecY-channel at long range, while substrates at the SecY-channel entrance feed back to regulate nucleotide exchange by SecA. This two-way communication suggests a new, unifying 'Brownian ratchet' mechanism, whereby ATP binding and hydrolysis bias the direction of polypeptide diffusion. The model represents a solution to the problem of transporting inherently variable substrates such as polypeptides, and may underlie mechanisms of other motors that translocate proteins and nucleic acids.DOI: http://dx.doi.org/10.7554/eLife.15598.001

Alignment of the protein substrate hairpin along the SecA two-helix finger primes protein transport in Escherichia coli

“…In addition, protein translocation in this system appears to use both SecA-dependent pushing and Brownian motion-dependent sliding to achieve efficient transport (33). Since the mobility and role of the THF subdomain in protein transport remain controversial (39,40), additional studies will be required to resolve this matter. Further studies can now address the biochemical and structural requirements needed to transition from the preinitiation to postinitiation states.…”

Bacterial Translocation Ratchets: Shared Physical Principles with Different Molecular Implementations

“…[98][99][100] Moreover, there is evidence that the bacterial Sec machinery employs and ATP-dependent Brownian ratchet mechanism for protein secretion. [101] Beyond bacterial secretion, the mechanism of translocation ratchet may be involved in import of bacteriophage DNA during the infection process. [102] In eukaryotic cells, precursor proteins are transported from the cytoplasm into mitochondria or the endoplasmatic reticulum.…”

“…[101] T4PS, type IV pilus system; T1SS, type I secretion system; T2SS, type II secretion system; T4SS, type IV secretion system. www.advancedsciencenews.com www.bioessays-journal.com hydrolysis and pmf whereas the later stages of translocation are powered by a translocation ratchet.…”

Dynamic action of the Sec machinery during initiation, protein translocation and termination revealed by single molecule fluorescence

“…Two principal models have been proposed: (1) a processive power-stroke in which a fixed length of substrate is transported for each ATP molecule hydrolysed by SecA (Erlandson et al, 2008;Zimmer et al, 2008); and (2) a Brownian ratchet model in which passive diffusive motion is conferred directionality by gating at the expense of ATP hydrolysis (Allen et al, 2016;Liang et al, 2009). …”

“…Recent studies favour at least some element of diffusion: previously, we proposed a "pure" stochastic model, in which the free energy available from ATP binding and hydrolysis at SecA drives a Brownian ratchet at the SecY LG (Allen et al, 2016), while others have suggested a hybrid processive/stochastic model, in which ATP hydrolysis generates a power stroke ('push') on the polypeptide, but the latter is allowed to diffuse through the pore ('slide') (Bauer et al, 2014). This marks a shift from deterministic models based on static structural snapshots to a stochastic view in which intrinsic dynamics of the complex are taken into account (Corey et al, 2016a).…”